Biology subjects

Pathmasiri, K. C.

Publications and source records attributed to Pathmasiri, K. C..

4 recordsLinked to original sources

bioRxiv2026

Immunohistochemistry-compatible gel-assisted mass spectrometry imaging

While lipids are fundamental to cellular signaling and structure, probing native lipid compositions and distributions across complex tissue architectures at single-cell resolution remains challenging due to inherent trade-offs in spatial resolution, sensitivity, and molecular coverage in current spatial lipidomics methods. Here, we report a multimodal imaging approach that seamlessly integrates gold-standard immunohistochemical labeling with gel-assisted mass spectrometry imaging to enable cell-type-specific, single-cell spatial lipidomics with modern instrumentation. Using intact brain tissue as a testbed, we demonstrate in situ measurement of cerebellar Purkinje cells and spatially mapped lipids across timepoints and cerebellar subregions within the pathological landscape of a neurodegenerative, lysosomal storage disorder. With single-cell lipidomic profiling, we delineated spatiotemporally distinct accumulation of specific glycosphingolipids and phospholipids within Purkinje cells and non-Purkinje cells in the diseased brain. Furthermore, unsupervised single-cell lipidomic clustering elucidated disease-progression-and subregion-dependent molecular divergence between healthy and neurodegenerative states within the cerebellum.

neuroscience↗

bioRxiv2026

Unveiling Lipid Dysregulation: Lipidomics of Mouse Brain and Isolated Myelin in Niemann - Pick Disease Type C1

Niemann-Pick Disease Type C1 (NPC1) is a fatal, neurodegenerative disorder, characterized by lysosomal lipid accumulation and dysmyelination. Previous studies have documented some lipid abnormalities in the null mouse (Npc1-/-) focused on the whole brain and liver. However, the specific lipidomic alterations in severely affected brain regions, such as cerebellum and isolated myelin remain understudied. We present a comprehensive LC-MS-based lipidomic analysis of the cerebellum and cortex of Npc1-/- mice during disease progression stages, along with the first comprehensive characterization of the myelin lipidome in NPC1 disease. Our results reveal that the cerebellum accumulates lipid species, including sphingolipids and glycerophospholipids progressively, while the cortex shows an overall decline in lipid levels, indicating region-specific lipid dysregulation. Notably, bis(monoacylglycero)phosphates and their precursors--including lysophosphatidylglycerol and hemibismonoacylglycerophosphate exhibit significant accumulation, with a preference for docosahexaenoic acid (DHA)-containing species. Despite known cholesterol storage defects in NPC1, we observed reduced free cholesterol levels in both regions, which we attribute to myelin loss. Myelin-specific lipidomics demonstrated extensive dysregulation, particularly in cortical myelin, including severe losses in sulfatides, ether-lipids, and acylcarnitine, alongside striking accumulation of hydroxy-ceramides. These findings identify novel lipid alterations in brain subregions and myelin, offering critical insight into the lipid perturbations under the loss of NPC1, and highlight lipid targets that may be crucial for therapeutic intervention and biomarker development.

biochemistry↗

bioRxiv2023

Gel-assisted mass spectrometry imaging

Compatible with label-free detection and quantification, mass spectrometry imaging (MSI) is a powerful tool for spatial investigation of biomolecules in intact specimens. Yet, the spatial resolution of MSI is limited by the methods physical and instrumental constraints, which often preclude it from single-cell and subcellular applications. By taking advantage of the reversible interaction of analytes with superabsorbent hydrogels, we developed a sample preparation and imaging workflow named Gel-Assisted Mass Spectrometry Imaging (GAMSI) to overcome these limits. With GAMSI, the spatial resolution of lipid and protein MALDI-MSI can be enhanced severalfold without changing the existing mass spectrometry hardware and analysis pipeline. This approach will further enhance the accessibility to (sub)cellular-scale MALDI-MSI-based spatial omics.

biochemistry↗

bioRxiv2023

DBDA matrix increases ion abundance of fatty acids and sulfatides in MALDI-TOF and mass spectrometry imaging studies

MALDI-TOF MS is a powerful tool to analyze biomolecules owing to its soft ionization nature and generally results in simple spectra of singly charged ions. Moreover, implementation of the technology in imaging mode provides a means to spatially map analytes in situ. Recently, a new matrix, DBDA (N1,N4-dibenzylidenebenzene-1,4-diamine) was reported to facilitate the ionization of free fatty acids in the negative ion mode. Building on this finding, we sought to implement DBDA for MALDI mass spectrometry imaging studies in brain tissue and successfully map oleic acid, palmitic acid, stearic acid, docosahexaenoic acid and arachidonic acid using mouse brain sections. Moreover, we hypothesized that DBDA would provide superior ionization for sulfatides, a class of sulfolipids, with multiple biological functions. Herein we also demonstrate that DBDA is ideal for MALDI mass spectrometry imaging of fatty acids and sulfatides in brain tissue sections. Additionally, we show enhanced ionization of sulfatides using DBDA compared to three different traditionally used MALDI matrices. Together these results provide new opportunities for studies to measure sulfatides by MALDI-TOF MS including in imaging modes.

biochemistry↗